Microdissection of the Prader-Willi syndrome chromosome region and identification of potential gene sequences

The Prader-Willi syndrome chromosome region on the long arm of human chromosome 15 was microdissected and microcloned from 20 GTG-banded metaphase chromosomes, and 5000 recombinant clones were obtained. Of these clones, 39% identify single-copy human DNA sequences, most of which map to the dissected chromosome region and are evolutionarily conserved in other species. Three of eleven clones studied in detail are deleted in several patients with Prader-Willi syndrome. The microclones will be useful for the physical characterization of the Prader-Willi syndrome chromosome region and the identification of the affected genes in this disease.     

Enzymic properties of intestinal aminopeptidase P: a new continuous assay

A continuous photometric assay of aminopeptidase P activity was developed which is based on a coupled enzymic assay with the substrate Gly-Pro-Pro-pNA and DPP IV as auxiliary enzyme. This assay was used to evaluate the kinetic parameters and inhibitory profile of intestinal brush border aminopeptidase P.     

NMR, CD and IR spectroscopies of a tridecanucleotide containing a no-base residue: coexistence of B and Z conformations.

The synthesis of the tridecadeoxynucleotide d(CGm5CGCGxACATGT), where x is the 1-cyano-2-deoxy-beta-D-erythropentofuranose, is described. The NMR, IR, CD studies at various salt concentrations and temperatures of this oligomer show that the B and Z conformations are simultaneously present in the same short DNA fragment. A single apurinic residue is sufficient for the coexistence of the B and Z helices on this oligomer.     

Ernährungsbiologie einer rhipidoglossen Kiemenschnecke

Zusammenfassung Die Arbeitsweise und die Leistungen der Rhipidoglossenradula von Theodoxus fluviatilis werden durch die kräftigen 4. Zwischenzähne und die Randbürstenzähne bestimmt. Die 4. Zwischenzähne lockern im wesentlichen die dem Fre\grund anhaftenden oder aufliegenden ein- bis wenigzelligen Algen (Diatomeen, chlorococcale und konjugate Grünalgen), die Randzähne fegen das gelockerte Nahrungsgut quantitativ zusammen. Fädige Grünalgen (z. B. Cladophorales) und Gewebeteile höherer Pflanzen werden nicht abgebissen bzw. abgeschabt.Die Diatomeen werden nur verdaut, wenn die Kieselschalen bereits bei der Nahrungsaufnahme mechanisch zerkleinert werden. Diese Zerkleinerung erfolgt allein auf einem Substrat mit rauher Oberfläche; sie wird durch die während des Bisses zwischen 4. Zwischenzähnen und Substrat auftretenden Reibungskräfte erzielt. Theodoxus wurde bei refiner Diatomeenernährung über mehrere Generationen gezüchtet. Tierisches Eiwei\ ist als Zusatzkost nicht erforderlich. Mit besonderen Hilfsma\nahmen kann Theodoxus im Laboratorium auch mit Cyanophyceen oder besonders mit Flagellaten (Chlamydomonas), die den Schnecken an den im Freiland besiedelten Standorten nicht zur Verfügung stehen, ernährt werden.Sämtliche Grünalgen mit stärkerer Cellulosewandung (Chlorococcales, Conjugatae) sind unverdaulich. Die Unverdaulichlichkeit beruht vermutlich auf einem Fehlen von Cellulasen im Verdauungstraktus. Die Zellmembranen und extrazellularen Scheiden der Cyanophyceen, die aus Hemicellulosen und Pektinen aufgebaut sind, werden im Magen aufgelöst. Theodoxus ist ein reiner Diatomeenfresser. Die ökologische Einnischung in die litorale Steinregion ist vorwiegend ernährungs-physiologisch begründet und erklärt das Vorkommen in Flie\gewassern und an Brandungsufern stehender Sü\gewässer sowie der Ostsee.     

Nuclear import and export of influenza virus nucleoprotein.

Influenza virus nucleoprotein (NP) shuttles between the nucleus and the cytoplasm. A nuclear localization signal (NLS) has been identified in NP at amino acids 327 to 345 (J. Davey et al., Cell 40:667-675, 1985). However, some NP mutants that lack this region still localize to the nucleus, suggesting an additional NLS in NP. We therefore investigated the nucleocytoplasmic transport of NP from influenza virus A/WSN/33 (H1N1). NP deletion constructs lacking the 38 N-terminal amino acids, as well as those lacking the 38 N-terminal amino acids and the previously identified NLS, localized to both the cytoplasm and the nucleus. Nuclear localization of a protein containing amino acids 1 to 38 of NP fused to LacZ proved that these 38 amino acids function as an NLS. Within this region, we identified two basic amino acids, Lys7 and Arg8, that are crucial for NP nuclear import. After being imported into the nucleus, the wild-type NP and the NP-LacZ fusion construct containing amino acids 1 to 38 of NP were both transported back to the cytoplasm, where they accumulated. These data indicate that NP has intrinsic structural features that allow nuclear import, nuclear export, and cytoplasmic accumulation in the absence of any other viral proteins. Further, the information required for nuclear import and export is located in the 38 N-terminal amino acids of NP, although other NP nuclear export signals may exist. Treatment of cells with a protein kinase C inhibitor increased the amounts of nuclear NP, whereas treatment of cells with a phosphorylation stimulator increased the amounts of cytoplasmic NP. These findings suggest a role of phosphorylation in nucleocytoplasmic transport of NP.     

Fiber analysis of human ventral and dorsal roots on the basis of different acetylcholinesterase activity]

Marked differences in the AChE activity of myelinated nerve fibers of ventral and dorsal roots could be established in human post mortem material. After a fixation time of 3 h and a critical incubation period of 24 h, in the mean 96% of the myelinated ventral root but only 4% of dorsal root fibers showed reaction product, detectable by the light microscope. The percentage of stained fibres varies, to some extent, in the different segments. Groups of very thin myelinated fibres within the ventral roots between the segments C-8 and L-3, showing a conspicuous high enzyme activity, are interpreted as pre-ganglionic sympathetic fibres; similar elements in the sacral ventral roots may represent parasympathetic fibres. The method of Karnovsky, applied under conditions established in this study, can be used for analysis of fibre types in a given human peripheral nerve.     

Nucleosome-associated protein kinases in murine erythroleukemia cells.

Chromatin subunits from murine erythroleukemia cells were prepared by a method which releases actively transcribing genes. Two casein kinase activities (CK₁ and CK₂) were isolated from these nucleosomes by gel nitration in 0.5 m NaCl. CK₁ (M_r ~ 200,000) and CK₂ (M_r ~ 35,000) were further purified by phosphocellulose chromatography and characterized with regard to several parameters which may regulate their activity in vivo. CK₁ has an NaCl optimum of 0.14 m, utilizes GTP as phosphate donor ~25% as efficiently as ATP, and phosphorylates a discrete group of high molecular weight nonhistone proteins in the unfractionated chromatin starting material. CK₂ has an NaCl optimum of 0.24 m, cannot utilize GTP, and modifies a different group of nonhistones. Both kinases are inhibited by concentrations of hemin (<50 μm) which efficiently induce globin gene expression in erythroleukemia cells. A histone kinase resolved during the gel filtration step is unaffected by hemin. An investigation of the mode of hemin inhibition reveals that CK₁ and CK₂ interact in different fashions with the inhibitor.